Dexamethasone-binding proteins in cytosol and nucleus of rat thymocytes. Purification of three receptor proteins.
Dexamethasone-binding proteins from the cytosol and the nucleus of rat thymocytes were analyzed by ion-exchange chromatography on DEAE-cellulose. Three dexamethasone-binding proteins were revealed in cytosol, one in the flow-through (DE-1) and two (DE-2 and DE-3) eluting from the column with 0.13 M and 0.23 M NH4Cl, respectively. In nuclear extracts only one receptor fraction, present in the flow-through, could be detected. By a combination of affinity chromatography on Cl-Sepharose to which dexamethasone 21-methanesulfonate was linked through a disulufide bond and DEAE-cellulose chromatography, three receptor proteins were highly purified from cytosol, with molecular weights of 45 000, 72 000 and 90 000 and one from nuclear extracts with molecular weight of 72 000. Antibodies to the 45 000-Mr and 90 000-Mr proteins were elicited in rabbits. The antibodies to the 45 000-Mr protein cross-react with the 90 000-Mr. Similarly, the antibodies to the 90 000-Mr protein cross-react with the 45 000-Mr protein. Antibodies to either of the two proteins immunoprecipitate 60--70% of the dexamethasone-binding activity of rat thymus cytosol. Immunoaffinity chromatography of cytosol and nucleosol on columns of Sepharose linked to the IgG against either the 45 000-Mr or the 90 000-Mr protein leads to binding of these proteins on the columns but not of the 72 000-Mr species. Two nuclear polypeptides with molecular weights of 36 000 and 38 000 remain attached to the immunoaffinity column; these polypeptides may represent degradation products of the cytoplasmic receptor upon entrance into the nucleus. Antibodies against two dexamethasone-binding proteins from rat liver cytosol immunoprecipitate the 45 000-Mr cytosol receptors from rat thymus.
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D003907:Dexamethasone / Q000378:metabolism*; D007106:Immune Sera; D007118:Immunoassay; D005779:Immunodiffusion; D008297:Male; D008970:Molecular Weight; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000302:isolation & purification* / Q000378:metabolism; D011987:Receptors, Steroid / Q000302:isolation & purification*; D013950:Thymus Gland / Q000378:metabolism*
D007106:Immune Sera; D011965:Receptors, Glucocorticoid; D011987:Receptors, Steroid; C028697:dexamethasone receptor; D003907:Dexamethasone